Despite years of intensive investigation, the genetic origin of antibody diversity remains an enigma. Amino acid sequence studies on myeloma proteins and antibodies will probably never fully answer this perplexing question, but as additional sequence work has been done, considerable constraints have been placed on both major theories of antibody formation. In all likelyhood, direct molecular probes of cellular DNA will, in the final analysis, lead to the answer. One major problem with much of the structural data available today is that it has been done on light chains. A serious difficulty in using chains concerns the presence of multiple V region subgroups (from nine in man to over 25 in the mouse, the most widely studied species). In the past four years we have mainly focused on the heavy chains of the immunoglobulins of man, mouse, cats and dogs. Three major pieces of information have emerged. (1) Heavy chains have far fewer subgroups in each of these species (probably three in man, two in mouse, and only one in cat and dog). (2) the structure of the V re@ion of the heavy chain has been remarkably perserved during evoluationary time (3) despite this extraordinary degree of structural conservation there are certain amino acids at particular positions in each of these species that are unique to the species ("phylogenetically associated residues"). This proposal concerns the extension of these observations and includes experiments which we hope will underscore the importance of these critical residues. We hope to (1) survey lower phylogenetic forms to trace the evolutionary history of the variable region gf the heavy chain, (2) search for additional phylegenetically associated residues in these lower species (3) study the nature of the antibody combining site to identical antigens in at least three species (4) study the interaction of heavy and light chain V region subgroups in man and then in lower forms.